Method for production of bio-ethanol using watermelon seeds

ABSTRACT

Provided is a method for production of bioethanol using watermelon seeds and, more particularly, a method for producing bioethanol with high production yield using watermelon seeds, including: sterilizing watermelon seeds, which are usually discarded as food waste from domestic houses, supermarkets, farm houses, etc., at 121° C. for 10 to 20 minutes under anaerobic conditions; finely grinding the sterilized watermelon seeds; adding glacial acetic acid to the ground seeds to remove linoleic acid therefrom; and inoculating the treated seeds free from linoleic acid with a strain for ethanol fermentation such as  Saccharomyces cerevisiae , followed by agitating at 25 to 35° C. and 100 to 300 rpm for 5 to 15 days, to conduct fermentation.

CROSS-REFERENCE TO RELATED APPLICATION

This application claims the benefit of Korean Patent Application No. 10-2010-0081535 filed on Aug. 23, 2010, in the Korean Patent and Trademark Office, the disclosures of which are incorporated herein in their entirety by reference.

BACKGROUND OF THE INVENTION

1. Field of the Invention

The present invention relates to a method of obtaining biofuels from watermelon seeds.

2. Description of the Related Art

Bioethanol is an eco-friendly alternative fuel extracted through fermentation of saccharide or cellulose fractions in plants. Such bioethanol increasingly draws more attention to its usefulness as an additive of gasoline products, as compared to existing biomethanol used as an additive for diesel products.

Since the 1970s, most studies into the development of bioethanol have been implemented in the United States and Brazil as the main producers. In Brazil, production of bioethanol using sugar cane has been promoted as one of national policy projects and about 30% of overall fuel consumption for vehicles was successfully replaced with bioethanol in 2004. For the United States, it was clarified in the President's Annual State of the Union address to Congress in 2008 that oil consumption would be reduced by 20%, instead, increasing the use of alternative energy such as bioethanol oil. In addition, other countries including, for example, Japan, China, other Asian nations, etc. are also moving ahead with policies to enlarge bioethanol production. As such, with increased demand and interest in bioethanol throughout the world, bioethanol production output is gradually increasing.

After the oil crisis in the 1970s, research and development into bioethanol begun as a part of alternative energy development and have mostly been executed with crop biomass obtained from corn and sugar cane and/or wood-based biomass obtained from wood resources, as raw materials. In fact, Brazil succeeded in fuel commercialization of bioethanol on the basis of abundant cultivation and supply of sugar cane. Meanwhile, plenty of commercial technologies to produce bioethanol utilizing abundant corn and/or wood resources have been accumulated in the United States.

However, with regard to crop biomass, ethical problems of utilizing invaluable food resources as fuel have recently been posed and, in addition, disadvantages in production of bioethanol using such crop biomass have also been encountered in terms of supply and demand of raw materials and/or price competitiveness, since international grain prices rapidly increased in early 2008.

Although resources for wood-based biomass have a merit of increasing ethanol production yield because they are plentiful in nature and content of cellulose and hemi-cellulose in the obtained biomass is at least 75%, the wood-based biomass has a physically and chemically rigid structure, as compared to crop biomass, thus entailing difficulties in chemical and/or enzyme treatment approaches for saccharification of cellulose and/or hemi-cellulose. Moreover, since wood-based biomass contains about 15 to 25% of lignin consisting of numerous hydrophobic aromatic compounds, high cost and complicated pre-treatment to remove such lignin fraction may be required. Furthermore, forest degradation encountered in biomass production also accelerates global warming and causes environmental damage, in turn entailing problems in environmental ethics such as violation of responsibility for eco-friendly fuel production.

Accordingly, in order to solve such ethical problems while overcoming energy (fuel) crisis or exhaustion that mankind is facing, there is a strong need to ensure novel and high efficiency biomass capable of replacing existing crop biomass or wood-based biomass.

SUMMARY

Accordingly, it is an aspect of the present invention to provide an eco-friendly method for production of bioethanol using watermelon seeds which are generally discarded as food waste.

Another aspect of the present invention is to provide a method for high efficiency production of bioethanol, by removing linoleic acid from watermelon seeds before ethanol fermentation thereof.

Yet another aspect of the present invention is to provide a method for producing bioethanol without ethical problems such as waste of food resources, as compared to utilization of existing crop biomass or wood-based biomass.

Still another aspect of the present invention is to provide yeast strains most effective for production of bioethanol using watermelon seeds and a method of maximally producing bioethanol by fermenting the yeast strains described above.

An embodiment of the present invention provides a method for production of bioethanol, including the steps of: sterilizing watermelon seeds at 121° C. for 10 to 20 minutes under anaerobic conditions; finely grinding the sterilized watermelon seeds; adding glacial acetic acid to the ground seeds to remove linoleic acid therefrom; and inoculating the treated seeds free from linoleic acid with a strain for ethanol fermentation.

According to the foregoing production method, after inoculating the prepared seeds free from linoleic acid with the strain for ethanol fermentation, the inoculated seeds may be subjected to fermentation while agitating at 25 to 35° C. and 100 to 300 rpm for 5 to 15 days.

According to the foregoing production method, the strain for ethanol fermentation used herein may be selected from Saccharomyces cerevisiae KCCM 1129, Saccharomyces cerevisiae KFCC 11352 or Saccharomyces sache KFCC 11513.

According to the foregoing production method, the strain for ethanol fermentation used herein may be prepared in a culture medium form by shaking incubation of Saccharomyces cerevisiae KCCM 1129 in a liquid medium including 5 g of peptone, 5 g of yeast extract and 5 g of glucose in 1000 ml of distilled water.

According to the foregoing production method, a relative ratio by weight of the prepared seeds free from linoleic acid to the strain for ethanol fermentation may range from 1:8 to 12.

According to the foregoing production method, a relative ratio by weight of the sterilized watermelon seeds to glacial acetic acid may range from 1:1 to 3.

According to the foregoing production method, during agitating after inoculating the prepared seeds free from linoleic acid with the strain for ethanol fermentation, a mineral solution including 1.0 g of (NH₄)₂SO₄, 1.0 g of KH₂PO₄, 1.0 g of K₂HPO₄, 0.2 g of MgSO₄.H₂O, 0.1 g of yeast extract, 10.0 mg of FeSO₄, 2.0 mg of CaCl₂, 2.0 mg of MgSO₄ and 2.0 mg of ZnSO₄ in 1 liter of water may be added to the inoculated watermelon seeds.

DETAILED DESCRIPTION

The present invention provides a method for production of bioethanol using watermelon seeds, which include: sterilizing watermelon seeds, which are usually discarded as food waste from domestic houses, supermarkets, farm houses, etc., at 121° C. for 10 to 20 minutes under anaerobic conditions; finely grinding the sterilized watermelon seeds; adding glacial acetic acid to the ground seeds to remove linoleic acid therefrom; and inoculating the prepared seeds free from linoleic acid with a strain for ethanol fermentation such as Saccharomyces cerevisiae, followed by agitating at 25 to 35° C. and 100 to 300 rpm for 5 to 15 days to conduct fermentation, so as to enable production of bioethanol with high production yield and considerably reduce release of hazardous substances such as carbon dioxide, formaldehyde, etc., thereby accomplishing eco-friendly and economically advantageous production of bioethanol without waste of food resources such as corn, sugar cane, or the like.

Hereinafter, embodiments of the present invention will be described in detail.

The present invention may produce bioethanol using watermelon seeds which are discarded as food waste. Although there is a difference in origins of watermelon, watermelon seeds generally contain 41.6% of carbohydrate such as sucrose, glucose, fructose, etc., 27.4% of fatty acid, 18.9% of protein, and the balance being other fractions such as mineral, vitamins, or the like. Among these, the carbohydrate fraction is converted into ethanol through anaerobic respiration of yeast having ethanol fermentation performance.

Firstly, watermelon seeds are subjected to sterilization at 121° C. for 10 to 20 minutes under anaerobic conditions. If the seeds are not suitably sterilized under such anaerobic conditions, fungi may be generated in large quantities and ethanol production yield may be reduced when the watermelon seeds are inoculated with yeast for ethanol fermentation during further processes.

Then, the sterilized watermelon seeds are finely ground using a grinder or the like. If the watermelon seeds are not finely ground, carbohydrates contained in the watermelon seeds may not efficiently react with the yeast or, during removal of linoleic acid using glacial acetic acid, reaction may not be sufficiently implemented.

Grinding procedures and/or conditions may be suitably adopted from conventional techniques, without being particularly limited thereto.

Then, the linoleic acid is removed from the ground watermelon seeds. For example, the ground watermelon seeds may be treated using glacial acetic acid then subjected to extraction of linoleic acid. Since linoleic acid molecules are stable due to a large number of carbon atoms therein, the yeast may not effectively use them as a carbon source so that the production of bioethanol may be inhibited. Although linoleic acid may be removed through alcohol extraction, glacial acetic acid is preferably used to remove linoleic acid. More preferably, 1 to 3-fold glacial acetic acid is added to a total weight of the ground watermelon seeds.

After removal of linoleic acid, the treated watermelon seeds may be inoculated with a strain for ethanol fermentation. The strains may include yeasts having ethanol fermentation performance, without being particularly limited thereto. In a case where watermelon seeds are used as biomass, the yeast may be any one selected from Saccharomyces cerevisiae KCCM 1129, Saccharomyces cerevisiae KFCC 11352 or Saccharomyces sache KFCC 11513. In consideration of production yield, Saccharomyces cerevisiae KCCM 1129 may be used.

After inoculating the strain for ethanol fermentation, the inoculated watermelon seeds are left for 5 to 15 days at 25 to 35° C. while being gently stirred at 100 to 300 rpm. If a fermentation temperature is outside the foregoing range, a rate for proliferation of strains may be undesirably decreased. It is advantageous that the strains sufficiently intake carbohydrate for at least 5 days. A proliferation period exceeding 15 days may not be suitable for anaerobic respiration.

Although an amount of the strain for ethanol fermentation used herein depends upon strain differences, a mixing ratio by weight of the prepared seeds free from linoleic acid to the strain for ethanol fermentation may range from 1:8 to 12. In particular, Saccharomyces cerevisiae KCCM 1129, Saccharomyces cerevisiae KFCC 11352 or Saccharomyces sache KFCC 11513, which is mixed with the watermelon seeds with the foregoing mixing ratio, may enable optimum production of ethanol. On the other hand, if the mixing ratio is beyond the foregoing range, a content of carbohydrate fraction in the watermelon seeds utilized by the strain is insufficient, in turn decreasing production yield.

The strain for ethanol fermentation is generally used for inoculation in a culture medium form prepared by shaking incubation of the strain in a liquid medium including 5 g of peptone, 5 g of yeast extract and 5 g of glucose in 1000 ml of distilled water. In addition, in order to improve a rate for proliferation of the strains, a mineral solution including 1.0 g of (NH₄)₂SO₄, 1.0 g of KH₂PO₄, 1.0 g of K₂HPO₄, 0.2 g of MgSO₄.H₂O, 0.1 g of Yeast extract, 10.0 mg of FeSO₄, 2.0 mg of CaCl₂, 2.0 mg of MgSO₄ and 2.0 mg of ZnSO₄ in 1 liter of water may be added to the inoculated watermelon seeds while stirring the watermelon seeds.

A resultant product, that is, ethanol obtained according to the foregoing processes may be collected by conventional separation methods (e.g., fractional distillation).

Hereinafter, preferred embodiments of the present invention will be explained to more concretely understand the present invention with reference to the following examples. However, it will be apparent to those skilled in the art that such embodiments are provided for illustrative purposes and various modifications and alterations may be possible without departing from the scope and spirit of the present invention, and such modifications and alterations are duly included in the present invention as defined by the appended claims.

EXAMPLE 1

Confirming the Possibility of Bioethanol Production from Watermelon Seeds

Saccharomyces sache KFCC 11513 and Saccharomyces cerevisiae KCCM 1129 were left at room temperature ranging from 25 to 30° C., thus being activated. After sterilizing watermelon seeds at 121° C. for 10 to 20 minutes under anaerobic conditions and grinding the same using a grinder, the ground watermelon seeds were divided into two groups, each of which weighed 20 g, using an electronic scale.

Both groups including the watermelon seeds were placed in Petri dishes and, one of the groups was inoculated with 100 g of a culture medium containing Saccharomyces sache KFCC 11513, while the other was inoculated with 100 g of another culture medium containing Saccharomyces cerevisiae KCCM 1129. Thereafter, plenty of water was added to each of the test groups, followed by sealing and leaving the same at room temperature.

After leaving the test groups about 7 days, ethanol generated in each dish was collected through fractional distillation and weighed. According to the foregoing procedures, 10.2 ml of ethanol was collected from the dish inoculated with Saccharomyces sache KFCC 11513 while 11.4 ml of ethanol was collected from the dish inoculated with Saccharomyces cerevisiae KCCM 1129. Consequently, it was confirmed that bioethanol may be produced using watermelon seeds.

EXAMPLE 2

Bioethanol Production after Removal of Linoleic Acid from Watermelon Seeds

It was found in Example 1 that bioethanol may be produced using watermelon seeds and the present example was performed to find how to maximize production yield of bioethanol. Among ingredients of the watermelon seed, linoleic acid was examined to determine whether it influences bioethanol production yield.

First, Saccharomyces cerevisiae KCCM 1129 were left at room temperature ranging from 25 to 30° C., thus being activated. The watermelon seeds were sterilized and ground according to the same procedures as described in Example 1. Next, a filter paper was spread in a funnel and the funnel was fixed above a beaker using a clamp and a stand. After placing 20 g of the ground watermelon seeds on the filter paper, 60 ml of glacial acetic acid was poured over the seeds using a glass rod.

After preparing two Petri dishes, the watermelon seeds free from linoleic acid were placed in one of the dishes while the same amount of watermelon seeds without removal of linoleic acid (as of the watermelon seeds free from linoleic acid) were placed in the other dish, followed by comparing ethanol production yields therebetween by the same procedures as described in Example 1. The comparison results exhibited that 12.3 ml of ethanol was formed in the dish containing the watermelon seeds free from linoleic acid while 11.0 ml of ethanol was obtained from the dish containing the watermelon seeds without removal of linoleic acid.

Consequently, it can be seen from the present example that ethanol production yield may be increased by removing linoleic acid from watermelon seeds before fermentation thereof with the yeast having ethanol fermentation performance.

EXAMPLE 3

Difference in Ethanol Production Yield Depending upon Strain Differences

Respective strains, that is, Saccharomyces cerevisiae KCCM 1129, Saccharomyces cerevisiae KFCC 11352 and Saccharomyces sache KFCC 11513, were left at room temperature ranging from 25 to 30° C., thus being activated. After sterilizing watermelon seeds at 121° C. for 10 to 20 minutes under anaerobic conditions and grinding the same using a grinder, the ground watermelon seeds were subjected to reaction with glacial acetic acid in order to extract linoleic acid therefrom. Next, the treated seeds were divided into three groups, each of which weighed 10 g, using an electronic scale.

Respective groups including the watermelon seeds were placed in three Petri dishes and were inoculated with 100 g of a culture medium containing Saccharomyces cerevisiae KCCM 1129, 100 g of a culture medium containing Saccharomyces cerevisiae KFCC 11352 and 100 g of a culture medium containing Saccharomyces sache KFCC 11513, respectively. Thereafter, 900 g of distilled water was added to each of the test groups, followed by fermentation at 30° C. and 180 rpm for 10 days.

As a result, it was found that 42.5 ml of ethanol was separated from the dish inoculated with Saccharomyces cerevisiae KCCM 1129, 36.5 ml of ethanol was separated from the dish inoculated with Saccharomyces cerevisiae KFCC 11352, and 38.2 ml of ethanol was separated from the dish inoculated with Saccharomyces sache KFCC 11513, respectively.

Consequently, it can be seen that the strain, Saccharomyces cerevisiae KCCM 1129, may be most suitable to produce ethanol from watermelon seeds as a raw material.

Moreover, a final amount of ethanol obtained using Saccharomyces cerevisiae KCCM 1129 is 4.25 ml/g. Assuming that watermelon production is approximately 37.2217 t/ha, each watermelon has an average weight of about 8 kg and contains seeds of about 20 g, watermelon seeds may be obtained in an amount of approximately 930,542.5 g/ha, thus resulting in bioethanol production of about 4,187.5 L/ha from the watermelon seeds. Consequently, it can be seen that watermelon seeds are more effective than corn which generally produces about 3,100 to 4,000 L/ha of bioethanol, in terms of bioethanol production.

EXAMPLE 4

Difference in Ethanol Production Yield Depending upon Ratio by Weight of Strain to Watermelon Seeds

Saccharomyces cerevisiae KCCM 1129 were left at room temperature ranging from 25 to 30° C., thus being activated. After sterilizing watermelon seeds at 121° C. for 10 to 20 minutes under anaerobic conditions and grinding the same using a grinder, the ground watermelon seeds were divided into three groups which weighed 5 g, 10 g and 20 g, respectively, using an electronic scale.

Respective groups including the watermelon seeds were placed in three Petri dishes and were inoculated with 100 g of a culture medium containing Saccharomyces cerevisiae KCCM 1129. Thereafter, 900 g of distilled water was added to each of the test groups, followed by fermentation at 30° C. and 180 rpm for 10 days.

As a result, it was found that 9.7 ml, 22.6 ml and 11.4 ml of ethanol were collected from these dishes containing 5 g, 10 g and 20 g of watermelon seeds, respectively.

These results exhibited that, when a ratio by weight of watermelon seeds to Saccharomyces cerevisiae KCCM 1129 is 1:10 during reaction therebetween, bioethanol production using watermelon seeds is most preferably accomplished. For the dish containing 5 g of watermelon seeds, an amount of carbohydrate used by the yeast strain is not sufficient to conduct fermentation. On the other hand, in the dish containing 20 g of watermelon seeds, chemically stable linoleic acid was considered to adversely affect activity of the yeast strain for ethanol production.

According to the present invention, it is possible to obtain eco-friendly biomass with high bioethanol production yield substantially equal to or more than those accomplished using corn, poplar wood, or the like. According to the present invention, carbon dioxide emissions may be maximally reduced by 90% or more, as compared to ethanol production through industrial processes. Moreover, release of toxic substances such as benzene, carbon monoxide, etc., during combustion may be remarkably decreased.

The present invention may provide an eco-friendly method for production of bioethanol by utilizing watermelon seeds which are discarded as food waste, enabling production of ethanol fuels useful for mankind while considerably reducing food waste.

Therefore, the present invention does not cause ethical problems such as wasting food resources, which is contrary to the utilization of existing crop biomass or wood-based biomass.

In addition, the present invention is based on findings regarding the most effective fermentation strains and conditions for production of bioethanol using watermelon seeds, thus enabling high efficiency production of bioethanol.

While the present invention has been described with reference to the preferred embodiments, it will be understood by those skilled in the related art that various modifications and variations may be made therein without departing from the scope of the present invention as defined by the appended claims. 

What is claimed is:
 1. A method for producing bioethanol, comprising: sterilizing watermelon seeds at 121° C. for 10 to 20 minutes under anaerobic conditions, grinding the sterilized watermelon seeds, adding glacial acetic acid to the ground seeds to remove linoleic acid therefrom; and inoculating the prepared seeds free from linoleic acid with a strain for ethanol fermentation.
 2. The method according to claim 1, wherein, after inoculating the prepared seeds free from linoleic acid with the strain for ethanol fermentation, the inoculated seeds are subjected to fermentation while agitating at 25 to 35° C. and 100 to 300 rpm for 5 to 15 days.
 3. The method according to claim 1, wherein the strain for ethanol fermentation is selected from Saccharomyces cerevisiae KCCM 1129, Saccharomyces cerevisiae KFCC 11352 or Saccharomyces sache KFCC
 11513. 4. The method according to claim 1, wherein the strain for ethanol fermentation is prepared in a culture medium form by shaking incubation of Saccharomyces cerevisiae KCCM 1129 in a liquid medium comprising 5 g of peptone, 5 g of Yeast extract and 5 g of glucose in 1000 ml of distilled water.
 5. The method according to claim 2, wherein a ratio by weight of the prepared seeds free from linoleic acid to the strain for ethanol fermentation ranges from 1:8 to
 12. 6. The method according to claim 1, wherein a ratio by weight of the sterilized watermelon seeds to glacial acetic acid ranges from 1:1 to
 3. 7. The method according to claim 2, wherein during agitating after inoculating the prepared seeds free from linoleic acid with the strain for ethanol fermentation, a mineral solution comprising 1.0 g of (NH₄)₂SO₄, 1.0 g of KH₂PO₄, 1.0 g of K₂HPO₄, 0.2 g of MgSO₄.H₂O, 0.1 g of yeast extract, 10.0 mg of FeSO₄, 2.0 mg of CaCl₂, 2.0 mg of MgSO₄ and 2.0 mg of ZnSO₄ in 1 liter of water is added to the inoculated watermelon seeds. 